Cxs are localized on the cell membrane of at least nine subtypes of APCs, namely, monocytes, macrophages, dendritic cells (DCs), including follicular dendritic cells (FDCs), Kupffer cells, B cells, astrocytes, microglia, neutrophils, and natural killer (NK) cells. Panx1 is also found on the eight subtypes of APCs ( Figure 1A ). As shown in Figure 1A , Cx43 is expressed in the aforementioned nine APC subtypes. Moreover, Cx37 is expressed on macrophages and neutrophils; Cx45 is present on the surface of DCs and microglial cells; Cx40 on the membrane of B cells and neutrophil cells; and Cx26 on astrocytes. In addition to those listed above, Cx36 and Cx32 are expressed on the microglial cell surface. Seven Cxs, namely, Cx30.3, Cx31.1, Cx32, Cx40, Cx43, Cx45, and Cx46, are present in T cells (4, 17, 18, 27–31). Unexpectedly, 8 of the 21 Cxs in the human gap junction protein family serve as components of synapses or participate in them. There are probably more Cx subtypes that remain to be identified in future research, as most previous studies focused on immune checkpoint molecules without conclusively evaluating Cxs in innate–adaptive immunity. Therefore, elucidating the interaction between HCs and IGLDCPs will offer mechanistic insights into the innate–adaptive immune response. In the following section, we have further detailed the mutual interaction between Cxs and IGLDCPs.

Cx43 interacts extensively with at least 20 IGLDCPS, either directly or indirectly, namely, CLMP, BCR, HepaCAM, CD4, CD8, CD19, CD25, CD3/CD28, CD31, CD39, CD40, CD46, CD47, CD60, CD69, CD73, CD80, CD86, IgG, and IgA ( Figure 1B ). Cx43 regulates T lymphocytes and DCs via IGLDCPs. Cx43/Cx40 maintains lymphocyte homeostasis and cytokine production, such as in the case of Cx43 HC inhibition, which suppresses IL-2 and IL-6 mRNA expression (32). Furthermore, the use of mimic peptides as an inhibitor of Cx43 HCs downregulated CD69 and CD25 activation in T cells, and led to IFN-γ by release by DC-stimulated NK cells (3). Similarly, CD3/CD28 induced ATP release by γδT cells, aided by HCs, resulting in cell activation (33, 34). Furthermore, Cx43 HCs in the plasma membrane of CD4⁺ T lymphocytes establish gap junction intracellular communication (GJIC) with macrophages to synthesize and secrete Igs and cytokines in immune regulation (19). Similarly, in DC–DC interaction, CD80, CD 86, and MHC class II are expressed (33, 35). Directly, Cx43 activates spleen cells and facilitates IgG production. Targeting Cx43 is a potential strategy to treat diseases associated with the antibody response (14). Cx43 regulates B lymphocyte activation directly, through BCR signaling, which involves migration and motility (7, 36).

The IGLDCPs regulate Cx43 HCs/GJs. The Ig-like domain in hepatocyte cell adhesion molecule (hepaCAM) stabilizes the Cx43 HCs on the cell surface. The hepaCAM gene was first described in hepatocellular carcinoma and was also discovered in the central nervous system (CNS); it is named GlialCAM, based on the site of its identification (37). HepaCAM is reportedly a member of the immunoglobulin superfamily (IgSF); it consists of an extracellular domain with two Ig loops, a transmembrane region, and a cytoplasmic tail, and functions in conjunction with Cx43 (38). In previous studies in U373 MG glioblastoma cells studies, it was found that hepaCAM expression redistributes Cx43, especially to the site of cell–cell contact, where co-localization of the two molecules is detected (38). Furthermore, altering the Ig-like domain of hepaCAM, especially the first extracellular IGLDCP reduces the co-localization of intercellular Cx43. Cx43 is shuttled back to the cytoplasm from the cell membrane, consequently decreasing its membrane-bound expression. In summary, the presence of IGLDCPs stabilizes Cx43 expression and promotes transport of a protein localized in the cytoplasm to the cell surface (22, 23). Additionally, CAR-like membrane protein (CLMP) regulates Cx43 and Cx45, and the absence of CLMP causes functional obstruction due to a lack of GJIC (24). Cxs and IGLDCPs are co-localized and interact at the immunological synapses (ISs). An increased intracellular Ca²⁺ level, which induces T cell activation and signal amplification, is facilitated by IS Cx43 HCs. Therefore, the formation of ISs is an important function structure, which allows us to understand how HCs/GJs and IGLDCP collaboratively modulate the precise roles in innate–adaptive immune responses.

Panx1 releases ATP from apoptotic cells to trigger an “eat me” signal (56) ( Figures 3A–C ). Key phagocytic inducers, ATP and UTP, have been confirmed to recruit apoptotic cells in vitro and in vivo. In contrast, “don’t eat-me” signals comprise CD31, CD46, and CD47 expression. These signals on healthy viable cells, which are capable of phosphatidylyserine (PtdSer) exposure under physiological conditions, may positively inhibit phagocytic uptake (26). These findings elucidate the mechanism governing HC–IGLDCP interaction in phagocytosis.

In the macrophage cell line J774, Cx43 RNAi showed impaired phagocytosis of the polystyrene-covered beads, and sheep erythrocytes opsonized by IgG (47); in contrast, in sheep erythrocytes with heterozygously or homozygously deleted Cx43, no changes were observed in phagocytosis (12, 57). Recently, Dosch et al. assessed Cx43 function in phagocytosis using Cx43 deletion and inhibition. It was determined that the inhibition of autocrine communication of Cx43-dependent ATP in macrophages improved sepsis outcome (33, 48, 49). Therefore, full expression of intact Cx43 is essential in regulating of the immune response through the directionality and rate of DC migration (58). Different cytokines regulate intercellular communication, facilitated by HCs/GJs in APCs, to execute purinergic signaling (3). This presents an interesting research opportunity for further investigation of Cx43-macrophage-phagocytosis. Apoptotic cells attract phagocytes by releasing chemotactic factors known as “find-me” signals (26).

Cumulative evidence shows that ATP triggers the inflammatory response. Cx HCs serve as a major pathway for the release of cytoplasmic ATP into the extracellular space. In granulocytes, Cxs enhance the inflammatory responses and promote cellular activation (33) ( Figure 4B ). For example, ATP released by Panx1 promotes the opening of Cx43 HCs (62) and is also involved in the innate immune response and inflammation (26, 63–70). In contrast, the blockage of Cx43 isoform HCs alleviates inflammation and enhances healing (2). In other inflammatory pathologies, Cx43 expression regulates monocyte–endothelial adhesion, with criteria for baseline adhesion set by Cx43-expressing monocytes (71). Similarly, elevated macrophage Cx43 HC activation and Panx1 expression inhibit pathogenesis (1, 49). Cx43 GJs transfer hypoxia-induced miR-192-5p, allowing cancer cells to acquire immune-resistant phenotypes (25). During inflammation in response to spinal cord injury, a decrease in the expression of Cx43 proteins shortens animal recovery time (33). ATP release has been inhibited using several Cx43 mimic peptides, thereby influencing the inflammatory process (72). Therefore, ATP integrated with the HC function promotes inflammation.

Open Cxs HCs in macrophages facilitate an effective immune response. GJs and HCs help spread toxicity into neighboring areas to augment viral/bacterial replication, and promote the spread of the inflammatory response by infectious agents, such as HIV (33),. Ig-like domains presenting T cell immunoglobulin molecules regulate inflammation and immune responses (16, 73). Single immunoglobulin IL-1R-related molecule (SIGIRR) is a specific inhibitor of IL-1R and toll-like receptor signals (74, 75).

In summary, Cxs HCs and Panx1 release ATP, which serves as an “eat me” signal; conversely, CD31, CD47, and CD46 function as “don’t eat-me” signals, which regulate phagocytosis in innate immunity.

The interaction between innate and adaptive immune response is defined as innate–adaptive immunity, which is important for antigen presentation. Cx43 contributes to trogocytosis ( Figures 4A, B ). The mechanism of innate control of adaptive immune responses involves multiple signaling pathways (16). We focused on how macrophages detect pathogens or injured cells. Trogocytosis is a process whereby lymphocytes extract surface molecules of APCs and express them on their own membranes (59, 60) ( Figure 4B ). However, the role of HCs and IGLDCPs in trogocytosis has not been examined adequately. Cx43-deleted macrophages are more proficient in T cell priming, implying an increased accumulation of antigens as these macrophages cannot transfer them to neighboring DCs, resulting in efficient presentation (57, 76–80). These findings delineate a potential mechanism by which HCs and IGLDCPs regulate antigen presentation.

GJs also have a pivotal function in DC activation and the amplification of antigen presentation, such as antigen transport, dendritic activation, and antigen cross-presentation (10, 33, 81–83). GJs-mediated antigen transfer between monocytes and CD8⁺DCs may serve as a simple and efficacious immunotherapy strategy for cancers, such as in the case of undifferentiated monocytes loaded with tumor antigen (20). Molecules containing Ig-like domains, such as pMHC-I and II, are involved in Cx43-dependent trogocytosis on the surface of acceptor cells (78). This is a valuable research direction to explore the underlying mechanism by which Cxs and IGLDCPs regulate antigen presentation via trogocytosis.

In addition to the roles of Cx43 in regulating macrophages and dendritic cells. HCs also mediate CD4⁺ T lymphocyte activation is critical in the transition of the innate immune response to adaptive immunity ( Figure 4C ). In a previous study, it was found that Cx43 in the IS delivers microRNAs from hypoxic melanoma cells to CTLs (25). Therefore, Cx43 stimulates T lymphocytes by the delivery of immune factors. Cx43 is involved in the formation of GJs in CD4⁺ T lymphocytes, Th0, Th1, and Th2, and macrophages; this pathway was found to be especially prominent in Cx43-Th1–macrophage interaction (19). This, in turn, suggests the potential capability of HCs in controlling IGLDCP activation. Cx43-GJs at the IS between DCs and CD4⁺ T cells promote T cell activation during antigen presentation (11), whereas the inhibition of GJs hindered DC-mediated T cell activation, reflected by lower T cell proliferation, CD69 expression, and IL-2 secretion.

Interestingly, in the absence of DCs, Cx43 GJ blockers did not affect the activation of CD4⁺ T cells triggered by anti-CD3/anti-CD28. Therefore, it was inferred that suppression of Cx43 inhibits Cx43 GJ assembly between DCs and T cells, resulting in T cell inactivation (84). In the DC–T cell IS, the blocking of Cx43 HCs/GJs (on either DCs or T cells) inhibited IFN-γ secretion and decreased the intracellular Ca²⁺ concentration, upon interaction of T cells with antigen-loaded DCs. These results strongly suggested that Cx43 HCs act in signaling amplification and T cell activation, by either releasing ATP or taking up of inositol triphosphate (IP₃) from DCs (25).

Cx43-GJs amplify antigens to activate T lymphocytes via the antigen cross-presentation pathway. In the immune synapse, Panx1, which releases ATP, controls Ca²⁺ entry to activate T cells; this happens by stimulating autocrine/paracrine receptors, such as P2X1 and P2X4 (85). Cx43-GJs between monocytes and CD8⁺ DCs transfer antigens (20), whereas Cx43 HCs activate CD4⁺ T cells (86). Together, Cx43-dependent trogocytosis of macrophages and dendritic cells promote antigen uptake, transfer, and presentation to activate innate–adaptive immunity. The GJ protein Cx43 induces B lymphocytes (8) to produce antibodies in plasma cells (61) ( Figure 4D ).

In cellular suppression mechanisms, naturally occurring Treg cells and helper T cells communicate via GJs to deliver cAMP to responder T cells, thereby inhibiting T cell proliferation and IL-2 synthesis (87–89) ( Figures 5A–C ). In a recent review, it has been reported how the cross-talk between Cxs and cAMP regulates cell-cycle progression, particularly in cancer cell populations (90). Furthermore, Cx43 expressed by thymic Treg cell progenitors supports Treg cell development. Conversely, Cx43 deletion decreased the number of functional Tregs and increased non-functional CD4⁺CD25⁺GITR⁺FOXp3^- T cells, which are incapable of producing inflammatory cytokines and inhibiting cancer cell progression (91). In human Treg cells, it has been shown that CD4-mediated activation involves elevation in the intracellular cAMP concentration. In contrast, the decrease in the cAMP level, caused by the application of adenylate cyclase (AC) inhibitor MDL12, resulted in the proliferation of Treg cells, in vitro and in vivo (87, 89) ( Figure 5B ). Consequently, it is inferred that Cx43 HCs may release cAMP; however, this needs to be studied further.

Cx43-GJs accumulate at the cytotoxic IS, enabling CTL-mediated melanoma cell killing (50). Additionally, Cx43 regulates the proliferation of CD4⁺CD25⁺ T lymphocytes and production of cytokines (92). Cx43-GJs regulate CD4⁺CD25⁺ Treg lymphocyte activation and inflammatory cytokine (IL-2 and IL-6) production in hypertensive inflammation in the spleen of rats (32, 92).

Recently, it was determined that tumor antigen-specific and tumor antigen-independent IgA transcytosis and antigen regulate ovarian cancer immunity. Tumor B cell-derived IgA binds to the polymeric immunoglobulin IgA receptors (pIgR) on ovarian cancer cells and reprograms myeloid cells against extracellular oncogenic drivers, such as EGFR and KRAS, which causes cell death. In particular, innate immunity triggered by antigen-independent IgA transcytosis is a novel strategy. IgA transcytosis through malignant epithelial cells causes tumor cells to encounter cytotoxic T cells, thereby hampering malignant progression; furthermore, the associated transcription changes result in suppression of the RAS pathway (21). In the ovarian cancer immunological response, IgA, B cells, and atypical B cells are observed (93). Transcytosis is a process in which molecules cross cellular barriers, which includes pinocytosis, endocytosis, and trafficking of vesicles to the opposite membrane (94).

In summary, Cx43-CD39/CD73-Treg-immunosuppression mediates adaptive immunity, specifically, IgA transcytosis, with tumor antigen-dependent and -independent mechanisms. It also regulates the establishment of immunity in ovarian cancer.